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The present study describes the Synthesis, characterization and biological activities of thiosemicarbazone ligand i.e. 3-Nitroacetophenonethiosemicarbazone (3NAT). The thiosemicarbazone ligand i.e. 3NAT has been synthesized by the condensation reaction of 3-Nitroacetophenone with thiosemicarbazide by conventional heating method as well as microwave irradiations method. The synthesized compounds have been characterized by elemental analysis, melting point determination, FTIR, UV-visible spectral analysis. The synthesized ligands have been screened in vitro for antibacterial activity against Escherichia coli, Staphylococcus aureus and Bacillus subtilis bacteria.

To research some medicinal importance of synthesized compounds.

All the chemicals and solvents used in the synthesis of ligands and transition metal complexes were of AR grade used as received. Thin layer chromatography was used to check purity of prepared compounds. IR spectra are recorded between the frequency 4000-500 cm-1 using KBr disc. Magnetic susceptibility of the synthesized transition metal complexes were measured on the vibrating sample magnetometer. Microwave synthesis was carried out in domestic microwave oven specially designed for green synthesis. Double Beam UV-Visible Spectrophotometerwas used for absorption measurement. All biological activities have been carried out with horizontal laminar.

A.    Preparations of Ligands

i. The Synthesis of 3-Nitroacetophenone Thiosemicarbazone (3NAT)

a. Microwave Irradiation Synthesis

A novel method for the preparation has been developed as per the principles of green chemistry, in which, either the reaction mixture was irradiated in a domestic microwave at 600 W for 2-10 minutes on alumina bed or reaction mixture in the solvent or the slurry of reaction mixture was exposed in a microwave reactor at 600 W maintaining different time intervals with respect to occasional and or definite inspection of TLC data. For the synthesis of 3NAT in accordance this novel synthesis method, water or water alcohol mixture of thiosemicarbazide (0.01mol) and 3-Nitroacetophenone (0.01mol) has been taken in Erlen Meyer flask capped with a funnel placed in a microwave oven and irradiated at 600 watt for 2-10 minutes. The reaction was monitored by TLC. After completion the reaction, the reaction mixture was allowed to attain room temperature and solid separated was filtered. The crude product was recrystalized from redistilled ethanol and dried under vacuum after filtration and separation. Purity of compound has been checked by TLC techniques in various solvents.

b. Conventional Thermal Method for Synthesis

In conventional thermal method, a hot ethanolic solution (25 mL) of 3-Nitroacetophenone (0.01 mol) has been mixed to a hot ethanolic refluxing solution (30 mL) of thiosemicarbazide (0.01 mol) in a 1: 1 molar ratio. The reaction mixture was refluxed in a water bath  for about 6-10 hours. Thin layer chromatography was used to check the progress of the reaction. After the concentration and cooling of the reaction mixture, product in crystallized form obtained. Purity of the compound was checked by TLC after a multiple washing and filtration.

The structures of ligands are shown in (Fig.II.B.13). A comparison between the thermal method and microwave method is given in (Table II.B.1).

ii. Synthesis of 4-Hydroxyaetophenone thiosemicarbazone (4HAT)

a. Microwave irradiation synthesis

In microwave irradiation preparation, water or water alcohol mixture of thiosemicarbazide (0.01mol) and 4-Hydroxyaetophenone(0.01mol) has been taken in Erlen Meyer flask capped with a funnel placed in a microwave oven and irradiated at 200 watt for 2-5 minutes. The reaction was monitored by TLC. After completion the reaction, the reaction mixture was allowed to attain room temperature and solid separated was filtered. The crude product was recrystalized from redistilled ethanol and dried under vacuum after filtration and separation. Purity of compound has been checked by TLC techniques in various solvents.

b.  Conventional Thermal method for synthesis

In conventional thermal method, a hot ethanolic solution (25 mL) of 4-Hydroxyaetophenone (0.01 mol) has been mixed to a hot ethanolic refluxing solution (30 mL) of thiosemicarbazide (0.01 mol) in a 1: 1 molar ratio. The contents have been refluxed for about 6-10 hours in a water bath. The reaction was monitored by TLC. The reaction was monitored by TLC. After the concentration and cooling of the reaction mixture product in crystallized form obtained. Purity of the compound was checked by TLC after a multiple washing and filtration.

The structures of ligands are shown in (Fig. II.B.17). A comparison between the thermal method and microwave method is given in (Table II.B.1)

TablE I
Physico-chemical Data of Thiosemicarbazones

(C.M. = Conventional method, M.M. = Microwave method)

Compound	Colour	Reaction Period		Yield (%)
		C.M. (Hrs)	M.M. (Min.)	C.M.	C.M.
3NAT	Brown	9.0	2.00	43	56
4HAT	Light Brown	8.0	2.30	53	64

Infrared Spectra

Fig.1.  IR spectra of 3NAT and 4HAT

Structure of 3NAT and 4HAT

iv. Biological Activities

The antibacterial activity of the compounds against E.coli, S.aureusand B.subtiliswere carried out using Muller Hinton Agar media (Hi media). The activity was carried out using paper disc method is represented in Table 2 which shows that both the 4NAT and 3NBT ligands have moderate antibacterial activities against these bacteria. Among the both ligands, 3NBT ligtand has been found out to be most effective against S.aureus bacteria showing maximum clarity of zones.

Fig.3 Biological avtivities

Table 2 Biological activities of Ligands

S. No.	Ligands	Zone of inhibition (in mm)
		E. coli	S. aureus	B. subtilis
1	4NAT	0.5	0.6	0.7
2	3NBT	0.5	8.0	0.6

The thiosemicarbazone ligand 3NAT and 4HAT were characterized by elemental analysis, spectral studies and magnetic moment measurements. On the basis of above study probable structure of the ligand is determined.. The antibacterial properties of the ligands were studied against E.coli, S.aureusand B.subtilis bacteria. The result shows that all the ligands have moderate antibacterial activities against these bacteria.

1. Parekh AK, Desai KR. Synthesis and antibacterial activity jof thiosemicarbazones. Indian journal of chemistry 2006; 45B: 1072-1075. 2. West DX, Padhye SB, Sonawane PB, Chikate RC. Copper(II) complexes of tridentate (O,N,S) thiosemicarbazones. Asian Journal of Chemistry Review 1990; 4(1): 125. 3. Agrawal PK, SinghalRenu, Arora SS, Agrawal Ajay. Synthesis and characterisation of Mn(II), Co(II) and Ni(II) complexes of semicarbazones and thiosemicarbazones of 2-pyridylglyoxal. International Journal for Chemical and Pharmaceutical Sciences 2015; 6(1): 30-35. 4. Ferrari MB, Bisceglie F, Fava GG, Pelosi G, Tarasconi P, Albertini R, Pinelli S. Synthesis, chacterisation and biological activity of two new polymeric copper(II) complexes with α-ketoglutaric acid thiosemicarbazones. J InorgBiochem 2002; 89(1): 36-44. 5. Singh K, Barwa MS, Tyagi P. Synthesis and characterisation of cobalt(II), nickel(II), copper(II) and zinc(II) complexes with Schiff base derived from 4-amino-3-mercapto-6-methyl-5-oxo-1,2,4,-triazine. Eur J Med Chem 2007; 42(3): 394-402. 6. Patil SD, Dawane BS, Kamble RD, Hese SV, Achaya AP, Kote JR, Gachhe RN. Synthesis and antimycobacterial activity of Cu(II) complexes containing thiosemicarbazones ligand. Der Pharmacia Sinica 2013; 4(2): 171-175. 7. Affan MA, Salam MA, Ahmad FB, White F, Ali HM. Organotin(IV) complexes of 2-hydroxyacetophenone-N(4)-cyclohexylthiosemicarbazone (H2dact):synthesis, spectral characterisation, crystal structure and biological studies. InorgChimActa 2012; 38: 219-225. 8. Bakheet TM, Doig AJ. Properties and identification of antibiotic drug targets. BMC Bioinformatics 2010; 11: 195. 9. Shim Jaesool, Jyothi NR, Mohammad Farook NA. Biological applications of thiosemicarbazones and their metal complexes. Asian Journal of Chemistry 2013; 25(10): 5838-5840. 10. Kumar Devesh, Singh VK.Application of metal complexes of schiff base with special reference to thiosemicarbazone: a review. Journal of Drug Discovery and Therapeutics2014; 2(13): 24-32. 11. French FA, Blanz EJ. The carcinostatic activity of α-(N-heterocyliccarboxaldehyde thiosemicarbazones II.3-hydroxypyridine-2-carboxaldehyde thiosemicarbazone. Cancer Res 1965; 25(9): 1454-8. 12. Kowol CR, Trondl R, Heffeter P et al. Impact of metal co-ordination on cytotoxicity of 3-aminopyridine-2-carboxaldehyde thiosemicarbazones (triapine) and no vel insights in to terminal dimethylation. J Med Chem 2009; 52(16): 5032-5043. 13. Mendes IC, Soares MA, Santos Dos RG, Pinheiro C, Beraldo H. Gallium (III) complexes of 2-pyridineformamide thiosemicarbazones: cytotoxic activity against malignant glioblastoma. Eur J Med Chem 2009; 44: 1870-7. 14. Kolesar JM, Schelman WR, Geiger PG et al. Electron paramagnetic resonance study of peripheral blood mononuclear cells from patients with refractory solid tumors treated with triapine. J InorgBiochem 2008; 102: 693-8. 15. Richardson DR, Sharpe PC, Lovejoy DB et al. Dipyridyl thiosemicarbazones chelators with potent and selective antitumor activity from iron complexes with redox activity. J Med Chem 2006; 49: 6510-21. 16. Chaston TB, Lovejoy DB, Watts RN, Richardson DR. Examination of the antiproliferative activity of iron chelators: multiple cellular targets and the different mechanism of action of triapine compared with desferrioxamine and the potent pyridoxal isonicotinoylhydrazone analogue 311. Clin Cancer Res 2003; 9: 402-14. 17.Mendes IC, Moreira JP, Speziali NL, Mangrich AS, Tskahashi JA, Beraldo H. N(4)- tolyl-2-benzoylpyridine thiosemicarbazones and their copper (II) complexes with significant antifungal activity. Crystal structure of N(4)-para-tolyl-2-benzolpyridine thiosemicarbazone. jBrazChemSoc 2006; 17(8): 1571-1577. 18. Ali S, Draksha. Synthesis, Characterization and Biological Evaluation of Some Cobalt(II), Nickel(II) and Copper(II) Complexes of 4[N(2’,4’ Dichlorobenzalidene)Amino]Thiosemicarbazone and 4[N(2’,4’Dinitrobenzalidene) Amino] Thiosemicarbazone. Asian J. Research Chem., 2011; 4(6): 976-983. 19. Fatondji HR, Gbaguidi F, Kpoviessi S, Hannaert V, Quetin-Leclercq J, Poupaert J, Accrombessi GC, Bero J, Moudachirou M. Synthesis, characterization and trypanocidal activity of some aromatic thiosemicarbazones and their 1,3,4- thiadiazolines derivatives. African Journal of Pure and Applied Chemistry 2011; 5(1): 59-64. 20. Fatondji HR, Kpoviessi S, Gbaguidi F, Bero J, Hannaert V, Moudachirou M, Quetin-Leclercq J, Poupaert J, AccrombessiGC. Structure activity relationship study of thiosemicarbazones on an African trypanosome: Trypanosoma bruceibrucei. Med Chem Res 2013; 22: 2151-2162. 21. Shipman C, Smith SH, Drach JC, Klaymanl DL. Thiosemicarbazones of 2-acetylpyridine, 2-acetylquinoline, 1-acetylisoquinoline and related compounds as inhibitors of herpes simplex virus in vitro and in a cutaneous herpes guinea pig model. Antiviral Reasearch 1986; 6: 197-222. 22. Gokhale N, Jain S. Synthesis and evaluation of novel thiosemicarbazones derivatives as anticancer agent. IJPSR 2015; 6(4): 1792-1804. 23. Yadav N, Bhojak N. Microwave assisted synthesis, spectral and antibacterial investigation on Co(II) complexes with amide ligands. The Int J EngSci 2013; 2(2): 166-168. 24. Silverstein RM, Bassler GC, Morrill TC. Spectroscopic identification of organic compounds, Johan Wiley and Sons: New York, 1991.